This proposal examines the contribution of receptor trafficking in inhibitory interneurons for establishing glomerulus-specific neuromodulation during olfactory. Current models of neuromodulation in olfaction emphasize the activation of specific classes of neurons that express a specific and dedicated class of neuromodulatory receptor, for example a single serotonin receptor. In such models, local interneurons in the first stages of olfactory processing may differentially innervate distinct olfactory glomeruli depending on the modulatory receptors that they express. Here, we will challenge this view and develop tools to demonstrate that odor specific modulation may occur not solely by targeting distinct cell classes, but also by the dendrite specific modulation of local interneurons that span multiple olfactory glomeruli. Specifically, we will develop tools to tag and track each serotonin receptor type in identified classes of local interneurons in the Drosophila antennal lobe, the first olfactory relay. Additionally, we will tag and label serotonin receptors in projection neurons that span multiple olfactory neuropil to determine how serotonin receptors are trafficked across olfactory neuropil. The Drosophila antennal lobe is an excellent model system for addressing these issues as its organization is highly similar to the mammalian olfactory bulb, and the smaller number of cells in the antennal lobe render it easier to study and characterize. Receptor tagging will be achieved be editing the fly genome to express modified serotonin receptors that are fused with the 11th fragment of the green fluorescent protein molecule, GFP. A genetic binary expression system can then be used to express the remaining portion of the split-GFP molecule to selectively label serotonin receptors in a specific cell class or individual neuron. We combine these tools with expansion microcopy to examine the co-localization of serotonin receptors with presynaptic and postsynaptic release sites within local interneuron and projection neurons populations. Finally, we will generate a version of these tools using a split mCherry fluorescent molecule approach so that the co-localization of serotonin receptor classes can be examined when local interneurons express multiple classes of these receptors. These experiments will lay the anatomical ground work for subsequent studies examining functional branch-specific neuromodulation of wide-field local interneurons in early olfactory processing.